XAF1 prevents hyperproduction of type I interferon upon viral infection by targeting IRF7.

Interferon regulatory factor (IRF) 3 and IRF7 are master regulators of type I interferon (IFN-I)-dependent antiviral innate immunity. Upon viral infection, a positive feedback loop is formed, wherein IRF7 promotes further induction of IFN-I in the later stage. Thus, it is critical to maintain a suitably low level of IRF7 to avoid the hyperproduction of IFN-I. In this study, we find that early expression of IFN-I-dependent STAT1 promotes the expression of XAF1 and that XAF1 is associated specifically with IRF7 and inhibits the activity of XIAP. XAF1-knockout and XIAP-transgenic mice display resistance to viral infection, and this resistance is accompanied by increases in IFN-I production and IRF7 stability. Mechanistically, we find that the XAF1-XIAP axis controls the activity of KLHL22, an adaptor of the BTB-CUL3-RBX1 E3 ligase complex through a ubiquitin-dependent pathway. CUL3-KLHL22 directly targets IRF7 and catalyzes its K48-linked ubiquitination and proteasomal degradation. These findings reveal unexpected functions of the XAF1-XIAP axis and KLHL22 in the regulation of IRF7 stability and highlight an important target for antiviral innate immunity.

A visualized ratiometric fluorescence sensing system for copper ions based on gold nanoclusters/perovskite quantum dot@SiO2 nanocomposites.

Excessive copper ions (Cu2+) cause serious environmental pollution and even endanger the health of organisms. Fluorescence chemosensing materials are widely used in the detection of metal ions due to their simple operation and high sensitivity. In this study, SiO2-encapsulated single perovskite quantum dot (PQD@SiO2) core-shell nanostructures which show strong, stable, and green fluorescence are synthesized and composited with gold nanoclusters (AuNCs) which show Cu2+-sensitive and red light-emitting fluorescence to obtain a visualized ratiometric fluorescence sensor (AuNCs/PQD@SiO2) for the detection of Cu2+. In the visualized detection of Cu2+, the green fluorescence emitted from the ion-insensitive PQD@SiO2 component is used as a reference signal and the red fluorescence emitted by ion-sensitive AuNC component is adopted as a sensing signal. In the presence of Cu2+, the red fluorescence is quenched whereas the green fluorescence remains stable, which results in a visualized fluorescence color change from orange-red to yellow and finally to green with increasing Cu2+ concentration. The significant change in the fluorescence color of AuNCs/PQD@SiO2 in response to Cu2+ enables a rapid, sensitive, and visualized detection of Cu2+. Further accurate and sensitive ratiometric fluorescence analysis of Cu2+ can be accomplished by measuring the ratio of fluorescence intensities at 643 and 520 nm (I643/I520) at a certain Cu2+ level. The developed AuNCs/PQD@SiO2-based sensor has been validated by its satisfactory application in the detection of Cu2+ in human serum and environmental water samples.

Colorimetric immunosensor based on glassy carbon microspheres test strips for the detection of prostate-specific antigen.

Micro-sized glassy carbon microspheres (GCMs, typically 3 µm in diameter) instead of nano-sized gold nanoparticles (AuNPs, typically 20 nm in diameter) were for the first time used as signal markers for the quantitative detection of antigen such as prostate-specific antigen (PSA). After being treated with concentrated HNO3, GCMs bear carboxyl groups at their surfaces, which enables antibodies to be conjugated with GCMs to yield new type of micro-sized material-based colorimetric probes used for immunochromatographic test strips (ICTSs). The captured black GCMs (with strong and wide-band light absorption) on the T-line of ICTS were used both for qualitative and quantitative determination of PSA. In the case of quantitative determination, a lab-assembled optical strip reader system was used to measure the reflected LED light intensity at 550 nm. The sensing performances of the developed GCM-based ICTSs, such as sensitivity, selectivity, reproducibility, stability, and applicability, were investigated in detail. The developed GCM-based ICTSs can have much higher (3 times) detection sensitivity than AuNP-based ICTSs, showing promising applications in sensitive immunoassay.

Multiomics analyses reveal a critical role of selenium in controlling T cell differentiation in Crohn's disease.

Inflammatory bowel disease (IBD) mainly includes Crohn's disease (CD) and ulcerative colitis (UC). Immune disorders play an essential role in the pathogenesis of these two IBDs, but the differences in the immune microenvironment of the colon and their underlying mechanisms remain poorly investigated. Here we examined the immunological features and metabolic microenvironment of untreated individuals with IBD by multiomics analyses. Modulation of CD-specific metabolites, particularly reduced selenium, can obviously shape type 1 T helper (Th1) cell differentiation, which is specifically enriched in CD. Selenium supplementation suppressed the symptoms and onset of CD and Th1 cell differentiation via selenoprotein W (SELW)-mediated cellular reactive oxygen species scavenging. SELW promoted purine salvage pathways and inhibited one-carbon metabolism by recruiting an E3 ubiquitin ligase, tripartite motif-containing protein 21, which controlled the stability of serine hydroxymethyltransferase 2. Our work highlights selenium as an essential regulator of T cell responses and potential therapeutic targets in CD.

Substrate-specific recognition of IKKs mediated by USP16 facilitates autoimmune inflammation.

The classic NF-κB pathway plays crucial roles in various immune responses and inflammatory diseases. Its key kinase, IKKß, participates in a variety of pathological and physiological processes by selectively recognizing its downstream substrates, including p105, p65, and IκBα, but the specific mechanisms of these substrates are unclear. Hyperactivation of one of the substrates, p105, is closely related to the onset of inflammatory bowel disease (IBD) in Nfkb1-deficient mice. In this study, we found that IKKß ubiquitination on lysine-238 was substantially increased during inflammation. Using mass spectrometry, we identified USP16 as an essential regulator of the IKKß ubiquitination level that selectively affected p105 phosphorylation without directly affecting p65 or IκBα phosphorylation. Furthermore, USP16 was highly expressed in colon macrophages in patients with IBD, and myeloid-conditional USP16-knockout mice exhibited reduced IBD severity. Our study provides a new theoretical basis for IBD pathogenesis and targeted precision intervention therapy.

Single-nucleotide methylation specifically represses type I interferon in antiviral innate immunity.

Frequent outbreaks of viruses have caused a serious threat to public health. Previous evidence has revealed that DNA methylation is correlated with viral infections, but its role in innate immunity remains poorly investigated. Additionally, DNA methylation inhibitors promote IFN-I by upregulating endogenous retrovirus; however, studies of intrinsically demethylated tumors do not support this conclusion. This study found that Uhrf1 deficiency in myeloid cells significantly upregulated Ifnb expression, increasing resistance to viral infection. We performed whole-genome bisulfite sequencing and found that a single-nucleotide methylation site in the Ifnb promoter region disrupted IRF3 recruitment. We used site-specific mutant knock-in mice and a region-specific demethylation tool to confirm that this methylated site plays a critical role in regulating Ifnb expression and antiviral responses. These findings provide essential insight into DNA methylation in the regulation of the innate antiviral immune response.

Stress-Induced Metabolic Disorder in Peripheral CD4+ T Cells Leads to Anxiety-like Behavior.

Physical or mental stress leads to neuroplasticity in the brain and increases the risk of depression and anxiety. Stress exposure causes the dysfunction of peripheral T lymphocytes. However, the pathological role and underlying regulatory mechanism of peripheral T lymphocytes in mood disorders have not been well established. Here, we show that the lack of CD4+ T cells protects mice from stress-induced anxiety-like behavior. Physical stress-induced leukotriene B4 triggers severe mitochondrial fission in CD4+ T cells, which further leads to a variety of behavioral abnormalities including anxiety, depression, and social disorders. Metabolomic profiles and single-cell transcriptome reveal that CD4+ T cell-derived xanthine acts on oligodendrocytes in the left amygdala via adenosine receptor A1. Mitochondrial fission promotes the de novo synthesis of purine via interferon regulatory factor 1 accumulation in CD4+ T cells. Our study implicates a critical link between a purine metabolic disorder in CD4+ T cells and stress-driven anxiety-like behavior.

A multilayer-graphene nanosheet film deposited on a ceramic substrate without a catalyst for constructing an electrochemiluminescence imaging platform.

Chemically and electrochemically stable conducting films are very desirable in the electrochemical industry and electrochemical sensing. In this work, ethanol was used as the carbon source to synthesize a multilayer-graphene nanosheet (MLGNS) film on ceramic substrates by a catalyst-free chemical vapor deposition (CVD) method at 900 °C and under ambient pressure. The developed CVD method is simple, economical and safe and avoids damage to the graphene nanosheet film during its transfer from the metal substrate to the non-metal substrate. The synthesized MLGNS film was well characterized by various techniques, including scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), and Raman spectroscopy. The prepared MLGNS film has good chemical and electrochemical stability and satisfactory electrical conductivity thus can be used as a new type of electrode material. The MLGNS film on the ceramic substrate has been fabricated into an electrochemiluminescence (ECL) imaging platform to investigate the oxygen reduction reaction (ORR) and evaluate the activities of ORR catalysts, such as PtNPs. The established MLGNS film-based ECL imaging platform may have promising applications in the study of catalysts for fuel cells, high throughput immunoassay in the clinic, and fast screening of anti-cancer drugs.

USP16-mediated deubiquitination of calcineurin A controls peripheral T cell maintenance.

Calcineurin acts as a calcium-activated phosphatase that dephosphorylates various substrates, including members of the nuclear factor of activated T cells (NFAT) family, to trigger their nuclear translocation and transcriptional activity. However, the detailed mechanism regulating the recruitment of NFATs to calcineurin remains poorly understood. Here, we report that calcineurin A (CNA), encoded by PPP3CB or PPP3CC, is constitutively ubiquitinated on lysine 327, and this polyubiquitin chain is rapidly removed by ubiquitin carboxyl-terminal hydrolase 16 (USP16) in response to intracellular calcium stimulation. The K29-linked ubiquitination of CNA impairs NFAT recruitment and transcription of NFAT-targeted genes. USP16 deficiency prevents calcium-triggered deubiquitination of CNA in a manner consistent with defective maintenance and proliferation of peripheral T cells. T cell-specific USP16 knockout mice exhibit reduced severity of experimental autoimmune encephalitis and inflammatory bowel disease. Our data reveal the physiological function of CNA ubiquitination and its deubiquitinase USP16 in peripheral T cells. Notably, our results highlight a critical mechanism for the regulation of calcineurin activity and a novel immunosuppressive drug target for the treatment of autoimmune diseases.

CRL4DCAF2 is required for mature T-cell expansion via Aurora B-regulated proteasome activity.

The proliferation of T cells in peripheral lymphoid tissues requires T cell receptor (TCR)-mediated cell cycle entry. However, the underlying mechanism regulating cell cycle progression in mature T cells is incompletely understood. Here, we have identified an E3 ubiquitin ligase, CRL4DCAF2, as a critical mediator controlling M phase exit in activated T cells. DCAF2 expression is induced upon TCR stimulation and its deficiency attenuates T cell expansion. Additionally, DCAF2 T cell-specific knockout mice display impaired peripheral T cell maintenance and reduced severity of various autoimmune diseases. Continuous H4K20me1 modification caused by DCAF2 deficiency inhibits the induction of Aurkb expression, which regulates 26S proteasome activity during G2/M phase. CRL4DCAF2 deficiency causes M phase arrest through proteasome-dependent mechanisms in peripheral T cells. Our findings establish DCAF2 as a novel target for T cell-mediated autoimmunity or inflammatory diseases.

CRL4DCAF2 negatively regulates IL-23 production in dendritic cells and limits the development of psoriasis.

The E3 ligase CRL4DCAF2 is believed to be a pivotal regulator of the cell cycle and is required for mitotic and S phase progression. The NEDD8-targeting drug MLN4924, which inactivates cullin ring-finger ubiquitin ligases (CRLs), has been examined in clinical trials for various types of lymphoma and acute myeloid leukemia. However, the essential role of CRL4DCAF2 in primary myeloid cells remains poorly understood. MLN4924 treatment, which mimics DCAF2 depletion, also promotes the severity of mouse psoriasis models, consistent with the effects of reduced DCAF2 expression in various autoimmune diseases. Using transcriptomic and immunological approaches, we showed that CRL4DCAF2 in dendritic cells (DCs) regulates the proteolytic fate of NIK and negatively regulates IL-23 production. CRL4DCAF2 promoted the polyubiquitination and subsequent degradation of NIK independent of TRAF3 degradation. DCAF2 deficiency facilitated NIK accumulation and RelB nuclear translocation. DCAF2 DC-conditional knockout mice displayed increased sensitivity to autoimmune diseases. This study shows that CRL4DCAF2 is crucial for controlling NIK stability and highlights a unique mechanism that controls inflammatory diseases.

Mitochondrial dynamics controls anti-tumour innate immunity by regulating CHIP-IRF1 axis stability.

Macrophages, dendritic cells and other innate immune cells are involved in inflammation and host defense against infection. Metabolic shifts in mitochondrial dynamics may be involved in Toll-like receptor agonist-mediated inflammatory responses and immune cell polarization. However, whether the mitochondrial morphology in myeloid immune cells affects anti-tumor immunity is unclear. Here we show that FAM73b, a mitochondrial outer membrane protein, has a pivotal function in Toll-like receptor-regulated mitochondrial morphology switching from fusion to fission. Switching to mitochondrial fission via ablation of Fam73b (also known as Miga2) promotes IL-12 production. In tumor-associated macrophages, this switch results in T-cell activation and enhances anti-tumor immunity. We also show that the mitochondrial morphology affects Parkin expression and its recruitment to mitochondria. Parkin controls the stability of the downstream CHIP-IRF1 axis through proteolysis. Our findings identify mechanisms associated with mitochondrial dynamics that control anti-tumor immune responses and that are potential targets for cancer immunotherapy.